Cell-free synthesis of ornithine decarboxylase. Changes in mRNA activity in the liver of thioacetamide-treated rats

Ornithine decarboxylase (ODC)mRNA associated with free polysomes of rat liver was translated in a reticulocyte lysate cell-free system. Newly synthesized ODC protein was identified by specific immunoprecipitation, molecular size as determined by polyacrylamide gel electrophoresis with sodium dodecyl sulfate, and competition by excess unlabeled ODC in the immunoprecipitation. A single injection of thioacetamide was found to cause several fold increases in both immunotitratable ODC protein and polysomal ODC-mRNA activity, while it provoked a much larger increase in ODC activity in rat liver. The results indicate that the induction of hepatic ODC activity by thioacetamide treatment is due not only to an increase in the activity of polysomal ODC-mRNA but also to a translational and/or posttranslational control.     

A direct evidence for defect in glucose-6-phosphate transport system in hepatic microsomal membrane of glycogen storage disease type IB

Uptake of glucose-6-phosphate by microsomes of hepatocyte in rats, human controls and patients with glycogen storage disease type Ia and Ib was studied. In rat the uptake of glucose-6-phosphate increased rapidly and reached to a plateau, but mannose-6-phosphate was not accumulated. These findings indicate that a glucose-6-phosphate specific transport system exists in the microsomal membrane. In human controls and patients with glycogen storage disease type Ia the uptake of glucose-6-phosphate was clearly observed. On the other hand, no accumulation of it was detected in a patient with glycogen storage disease type Ib. These data provide a direct evidence of the defect in the glucose-6-phosphate transport system of hepatic microsomal membrane in glycogen storage disease type Ib.     

Loose association of ribulose 1,5-bisphosphate carboxylase/oxygenase with chloroplast thylakoid membranes

The intra-chloroplastic distribution of ribulose 1,5-bisphosphate carboxylase/oxygenase (RuBisCO) between thylakoid membranes and stroma was studied by determining the enzyme activities in the two fractions, obtained by the rapid centrifugation of hypotonically disrupted chloroplast preparations of spinach and pea leaf tissues. The membrane-associated form of RuBisCO was found to increase in proportion to the concentration of MgCl₂ in the disrupting medium; with 20 mM MgCl₂ approximately 20% of the total RuBisCO of spinach chloroplasts and 10% of that of pea chloroplasts became associated with thylakoid membranes. Once released from membranes in the absence of MgCl₂, addition of MgCl₂ did not cause reassociation of the enzyme. The inclusion of KCl in the hypotonic disruption buffer also caused the association of RuBisCO with membranes; however, up to 30 mM KCl, only minimal enzyme activities could be detected in the membranes, whereas above 40 mM KCl there was a sharp increase in the membrane-associated form of the enzyme.Higher concentrations of chloroplasts during the hypotonic disruption, as well as addition of purified preparations of RuBisCO to the hypotonic buffer, resulted in an increase of membrane-associated activity. Therefore, the association of the enzyme with thylakoid membranes appears to be dependent on the concentration of RuBisCO. P-glycerate kinase and aldolase also associated to the thylakoid membranes but NADP-linked glyceraldehyde-3-P dehydrogenase did not. The optimal conditions for enzyme association with the thylakoid membranes were examined; maximal association occurred at pH 8.0. The association was temperature-insensitive in the range of 4° to 25° C. RuBisCO associated with the thylakoid membranes could be gradually liberated to the soluble form upon shaking in a Vortex mixer at maximal speed, indicating that the association is loose.Abbreviations DTT dithiothreitol - RuBP ribulose 1,5-bisphosphate - RuBisCO ribulose 1,5-bisphosphate carboxylase/oxygenase - MES 2-(N-morpholino) ethane sulfonic acid     

Effect of sulpiride on serum growth hormone and prolactin concentrations following L-DOPA administration in man.

The effect of chronic administration of sulpiride on serum human growth hormone (hGH), prolactin and thyroid stimulating hormone (TSH) was examined in 6 normal subjects. Sulpiride was given orally at a dose of 300 mg (t.i.d.) for 30 days. Sulpiride raised serum prolactin levels in all subjects examined. In addition, sulpiride suppressed hGH release induced by L-dopa, although the basal hGH level was not changed. Sulpiride treatment appeared to antagonize partially the inhibitory effect of L-dopa on prolactin release. Following thyrotropin-releasing hormone (TRH) injection, the percent increment in prolactin levels from the baseline in sulpiride-treated subjects was less than in controls without sulpiride. In contrast, both the basal and TRH-stimulated TSH levels were not influenced by sulpiride. These observations suggest that sulpiride suppresses L-dopa-induced hGH release and stimulates prolactin release, presumably by acting against the dopaminergic mechanism either on the hypothalamus or on the pituitary. The decreased prolactin response to TRH after sulpiride treatment may indicate a diminished reserve capacity in pituitary prolactin release.     

Microtubular structures in a stable staphylococcal L-form.

Microtubular structures, which were demonstrated as straight, dense-walled cylinders attached to cell membranes, were found in a stable staphylococcal L-form grown in the absence of antibiotic.     

Modes of DNA replication primed with the oligomer of palindrome

What is the precise molecular mechanism of semi-conservative DNA replication? After the great efforts of the past 20 years, molecular biology has now established the discontinuous syntheses of daughter DNA on both of the parental strands. In order to explain this type of discontinuous replication, we introduce the concept of a palindromic primer.First we focus our attention on various oligomers (RNA or DNA) which appear usually or occasionally in the process of replication. Then we propose the palindromic nature of these oligomers so as to serve as the primer of DNA synthesis. This postulation gives a theoretical reasoning for the discontinuities of both new strands in the fork region of replication.Subsequently we consider Watson's concatemeric intermediate theory, proposed for the explanation of replicative synthesis of phage T7 DNA. By considering the contribution of some sequence-specific endonuclease(s), we suggest the existence of partial palindromic sequences of bases at the connecting region(s) in which the redundant ends of the respective phage DNA molecules are overlapping. Another theory on the replication of linear chromosomal DNA including the concept of the terminal palindromic sequence of bases is also analyzed from the viewpoint of palindromic primer. Further, some recent experimental approaches, especially on the origin(s) of DNA replication, are shown to favour the concept of a palindromic primer.     

A general characteristic of tumor mitochondria: Leakage of endogenous Mg2+ on incubation with uncoupler and resultant reduction of uncoupler-stimulated ATPase activity

Previous studies showed that stimulation of mouse mitochondrial ATPase activity of tumor cells, fetal liver, and adult brain by the uncoupler 2,4-dinitrophenol was markedly suppressed during incubation of the mitochondria with the uncoupler (J.-I. Hayashi et al., 1980, Biochem. Biophys. Res. Commun.92, 261–267). The present work showed the reason for this suppression. More than half the endogenous Mg²⁺ leaked from mitochondria of all tumor cells tested, and of fetal liver and adult brain during incubation with the uncoupler, while only about 30% of the endogenous Mg²⁺ leaked from mitochondria of other normal tissues. The effect of the uncoupler on Mg²⁺ leakage from liver mitochondria changed from the fetal to the adult type within about 30 min after birth. In hypotonic medium, normal liver mitochondria also lost more than half their total Mg²⁺ and concomitantly stimulation of their ATPase activity by uncoupler was considerably reduced. Exogenously added Mg²⁺ could reverse this reduced effect of the uncoupler on ATPase activity of mitochondria from normal tissues and tumor cells. These results show that the endogenous Mg²⁺ content of mitochondria directly affects the stimulation by uncoupler of ATPase activity of mitochondria from both normal tissues and tumor cells. Thus, mitochondria of all tumor cells tested, and of fetal liver and adult brain are leaky to Mg²⁺ during incubation with uncoupler and as a result of the leakage, the stimulatory effect of the uncoupler on their ATPase activity is greatly reduced.